Tobacco usage causes or facilitates chronic bronchitis and atherosclerosis of the blood arteries, common among smokers. It also causes a rare, but characteristic disease of the small arteries of the limbs, thromboangiitis obliterans (TAO). The bronchial epithelial cells (BEC) that line the bronchi, and the endothelial cells (EC) which line the blood vessels express a variety of acetylcholine receptors (nAChRs) sensitive to nicotine (Nic). Nic is an agonist for some of the nAChRs present on the BEC and EC (the alpha3, alpha4 and alpha7 types), and an antagonist for others (the alpha9 and alpha9/alpha10 nAChRs). We wish to test the hypothesis that the nAChRs expressed by the BEC and EC mediate local cholinergic signals which modulate important cellular functions; and that exposure to Nic affects those functions, perhaps causing or facilitating some of the pathology induced by tobacco usage in the respiratory and cardiovascular systems. During the current grant period, we have examined the presence and function of nAChRs of the alpha3 subtype in BEC and EC. In these proposed studies, we wish to examine the expression and function of the nAChRs expressed by EC and BEC, the alpha7 and alpha9 nAChRs, and determine the effect of Nic exposure on the cellular functions controlled by these nAChRs. We also want to examine whether the BEC and EC secrete an acetylcholine-binding protein (ACh-BP) able to bind and sequester ACh and Nic, thereby terminating their action. Finally, we want to investigate whether an abnormality in the synthesis or function of EC nAChRs or ACh-BP occurs in TAO patients. The specific aims will be: 1. To identify cellular functions controlled by activation of alpha7 and alpha9nAChRs in EC and BEC. Specifically, to examine the effect of stimulation or specific blockade of those nAChRs on cell proliferation, apoptosis and cell death; and on the synthesis and release of NO, and of cytokines. 2. To determine the effect of Nic exposure on the synthesis of the EC and BEC nAChRs; on the synthesis of anti-inflammatory and pro-inflammatory cytokines and chemokines; and on the synthesis of NO. 3. To determine whether EC and BEC synthesize an ACh-BP that modulates cholinergic signaling; and whether secretion of ACh-BP is modulated by nAChR activation. 4. To examine whether the synthesis or function of EC nAChRs, and possibly ACh-BP, is altered in TAO patients.